FIGURE 4.

Effects of Bt₂cAMP on the NF-κB-dependent transcription and IL-8. A, AKIP1 switches the role of PKA signaling in NF-κB-dependent luciferase expression. Left panel, 293 (closed columns) and MDA-MB231 cells (open columns) were treated with Bt₂cAMP (1 and 3 mm) for 12 h and TNFα (1 ng/ml) for an additional 6 h. Cells were lysed and subjected to luciferase assay and normalized with the Renilla luciferase activity as an internal control. The vertical axis indicates the relative luciferase activity in fold activation. Values are representative of triplicate experiments (mean ± S.D.). Right panel, 293 cells transfected with (open columns) or without FLAG-AKIP1 (closed columns) were treated with Bt₂cAMP and TNFα and subjected to luciferase assay. Values are representative of triplicate experiments (mean ± S.D.). The inset in the right panel indicates the expanded representation of results in lanes 1–3 (the vertical axis indicates the fold activation). B, effects of AKIP1 on the NF-κB-dependent expression of IL-8. 293 and MDA-MB231 cells were cultured in 24-well plates and incubated with Bt₂cAMP and TNFα. The concentration of IL-8 was measured by ELISA. The vertical axis indicates the secretion of IL-8 in the culture medium, and the values are representative of triplicate experiments (mean ± S.D.).